DESCRIPTION (Applicant's Abstract): The African trypanosome is a protozoan parasite that causes a fatal disease called sleeping sickness in humans and ngana in domestic livestock. It is also a highly valued experimental organism, whose study has contributed fundamental discoveries in biology. Sequencing of the nuclear genome of Trypanosoma brucei reference strain TREU927/4 GUTat10. 1 commenced in 1998 at both The Institute for Genomic Research (NIAID U01 AI43062) and Sanger Centre (Wellcome Trust). TIGR has since provided an invaluable scaffold for chromosome mapping in the form of approximately 11,000 BAC and P1 end sequences, and approximately 37,000 shotgun sequences for gene discovery. In 1999, we initiated the sequencing of chromosomes II, IV and VI, using a 'map-as-you-go' approach; and will shortly produce the first complete sequence of an African trypanosome megabase chromosome (chr II). The Sanger Centre has carried out megabase chromosome sequencing by whole chromosome shotgun and skim sequencing of mapped genomic clones. They expect to complete the sequence of chr I very soon and have secured funding to sequence the three largest chromosomes of T. brucei (IX, X and XI). The remaining chromosomes (III, V, VII and VIII) were provisionally adopted by TIGR. We now propose to (i) determine the 7 Mb sequence of the four remaining chromosomes using the same approach used successfully to sequence chr II, by combining existing genome resources with a map-as-you-go strategy based on the use of end sequences from BAC clones to select optimal clones for sequencing; (ii) analyze and annotate the sequence; and (iii) make the sequence and analysis data available to the research community. The complete genome sequence of T. brucei will provide invaluable information and benefits at many levels, including (1) identification of genes involved in basic functions of the eukaryotic cell, (2) easy, inexpensive and fast cloning of genes encoding proteins being actively studied in laboratories around the world, (3) immediate access to genes and their products for functional/structural studies, (4) prediction of metabolic pathways on the basis of candidate genes, and (5) identification of parasite-specific gene products by comparison with other genomes.